The present invention relates to a transdermal delivery system containing an active ingredient selected from the group consisting of peptides and proteins having an Sxe2x80x94S bond and mixture thereof. These systems incorporate further components that assist in stabilizing the active material, e.g., by preventing the inactivation thereof and facilitating the penetration thereof as active molecules through the skin layers.
It has been discovered in the present invention that oxidizing agents such as iodine, povidine-iodine, potassium permanganate, peroxides and silver protein enable the application of formulations containing proteins and peptides and especially insulin onto the skin.
Insulin is secreted from beta cells of pancreatic Langerhans islet in its active form. The human insulin is composed of two polypeptides, the A and B chain, usually of 21 and 30 amino acids residues, respectively, with a molecular mass of about 5800 Dalton. The peptides are interconnected by disulfide bonds of the cysteine residues at A7-B7, A20-B19 and A6-A11. Insulin exerts a wide variety of biological activities including controlling the uptake, utilization, and storage of cellular nutrients such as glucose, amino acids and fatty acids. The important target tissues of insulin are liver, muscle and fat but many other cell types are also influenced by this hormone. [Davis, S N; Granner,D K (1996) In: Goodman and Gilman""s The Pharmacological Basis of Therapeutics. ninth edition ed. (Eds: Hardman,J G et al.) McGraw-Hill, 1487-15171].
One of the main physiological roles of insulin is stimulation of glucose transport into muscle and adipose tissues. A defect in this system leads to the diabetes mellitus syndrome characterized by hyperglycemia, changes in the metabolism of carbohydrates, lipids and proteins, and by elevated incidence of vascular disorders. There are two main diabetes, insulin-dependent diabetes mellitus (IDDM) with an incidence of 1-43 per 100,000 inhabitants in the Western countries, and the non-insulin-dependent diabetes mellitus (NIDDM) whose incidence is between 100-800 per 100,000 inhabitants in the Western countries (The above reference).
Insulin is the main treatment of all IDDM and many NIDDM patients. Long-term treatment is predominantly based on subcutaneous administration of insulin formulations. There are long-, short- and intermediate-acting preparation which are used according to the special requirements of the patient. However, apart from the discomfort and troublesome feelings and the possibility of infection associated with daily injections along the entire lifetime (particularly with IDDM, formerly termed juvenile-onset diabetes mellitus), this kind of therapy has serious clinical problems mainly with the maintenance of the appropriate blood levels of the hormone resulting in non-physiological blood glucose levels and other complications. Although much effort has been made in developing insulin analogs [Brange,J; et al. (1990) Diabetes 13, 923-954] and genetic engineering methodologies [Sutherland, DER et al. (1989) Diabetes 38 Suppl 1, 46-54], there are no successful findings for solving the clinical problems associated with parenteral insulin injections.
One of the approaches aimed to cope with this difficulty was to deliver the hormone non-invasively, via transdermal route of administration. By this procedure, the annoyance and inconvenience of the parenteral injections can be avoided; moreover, much steadier blood hormone levels can be achieved due to prolonged delivery of the drug. Several low molecular weight drugs have been formulated and are being clinically used as transdermal preparations. However, apart from a few medicines, many drugs, particularly peptides and proteins, are not successfully formulated for transdermal delivery. In vitro experiments have shown that xcex1-melanocyte stimulating hormone analog can penetrate across human and mouse, but not in rat skin [Dawson, B V et al. (1990) J. Invest. Dermatol. 94, 432-435; Dawson, B V et al. (1988) Life. Sci. 43, 1111-1117] and that enkephalin can penetrate hairless mouse skin but in the presence of the enhancer n-decylmethyl sulfoxide and proteinase inhibitors [Choi,H K et al. (1990) Pharm. Res. 7, 1099-1106]. However, apart from one study with small number of mice which showed reduced levels of blood glucose after 4 hours of cutaneous application of insulin with enhancer [Liedtke,R K et al. (1990) Drug Res. 40, 880-883], no efficient in vivo transdermal penetration of peptides and proteins, by chemical means (e.g. enhancer or proteinase inhibitors) have been published.
Transdermal penetration of various peptides and proteins can be enhanced by iontophoresis using electrical current for delivering charged agents across the skin. Various peptides and small proteins including insulin, calcitonin, vasopressin, luteinizing hormone-releasing hormone, leuprolide, thyrotropin-releasing hormone and cholecystokinin were tested in in vitro iontophoresis assays and some of them also in in vivo systems [Heit,M C et al. (1993) J Pharm Sci 1993 82(3):240-243; Srinivasan,V et al. (1990) J. Pharm. Sci. 79, 588-591; Burnette,R R and Marrero,D (1986) J. Pharm. Sci. 75, 738-743; Banga,A K and Chien,Y W (1993): Pharm. Res. 10, 697-702; Mao,X M et al. (1995) Yao. Hsueh. Hsueh. Pao. 30, 302-306; Mao,X M et al. (1995) Yao. Hsueh. Hsueh. Pao. 30, 881-885; Meyer,B R et al. (1989) Am. J. Med. Sci. 297, 321-325]. Additional technique to facilitate transdermal delivery of insulin by ultrasound vibration, termed sonophoresis, was used in both in vitro and in vivo systems [Tachibana,K and Tachibana,S (1991): J. Pharm. Pharmacol. 43, 270-271; Tachibana,K (1992) Pharm. Res. 9, 952-954; Mitragotri,S et al. (1995) Science. 269(5225), 850-853]. Although transdermal penetration of insulin and other proteins and peptides was enhanced by the sonophoretic and iontophoretic techniques, these procedures require complicated and an uneasy way of operation. Furthermore, the safety of long-term, daily use of this technique was not confirmed. The fact that only one report, describing unsatisfactory results on the use of penetration enhancer in type II diabetic patients [Liedtke,R K et al. (1990): Drug Res. 40, 884-886] have been published, indicates the problematic issues of the above methods.
According to U.S. Ser. No. 09/424525 there is provided a transdermal delivery system for treating diabetes and insulin influenced pathologic systemic conditions comprising insulin and a pharmaceutically accepted oxidizing agent selected from the group consisting of permanganate and silver protein, said oxidating agent enabling and facilitating the penetration of said insulin through the skin layers and into the blood stream.
According to the present invention there is now provided a transdermal delivery system for treating diabetes and other pathologic systemic conditions, comprising an active ingredient selected from the group consisting of peptides and proteins having an Sxe2x80x94S bond and mixtures thereof and a pharmaceutically acceptable oxidizing agent selected from the group consisting of iodine, povidine-iodine and sources thereof, said system being essentially free of reducing agents and said oxidizing agent enabling and facilitating the penetration of said active ingredient through the skin layers and into the blood stream.
As will be discussed hereinafter it is believed that said oxidizing agent serves to oxidize reduced glutathione thereby preventing its functioning as an inactivating agent.
Assuming that this hypothesis is correct then also a component such as buthionine sulfoximine, which also prevents the formation of glutathione can be used in the present invention alone or in combination with an oxidizing agent to achieve the desired effect.
In preferred embodiments of the present invention said active ingredient is insulin.
In WO 90/00899 by Rothman there are described compositions for treating conditions of keratinous tissue in mammals including wounds, sebborhea, psoriasis, dandruff, acne, itching, allergic reactions, non-specific dermatitis eczematoid dermatitism chronic dermatitis, equine exuberant granuloma, decubitis ulcers, and canine cutaneous granulomas which compositions include an activated protein component a compatible reducing agent and an oxidizing agent. While said compositions include a protein and an oxidizing agent and are applied to the skin the types of treatments disclosed therein are completely different from the transdermal delivery system of the present invention and as demonstrated in the comparative hereinafter the compositions of said publication are incapable of achieving a transdermal effect. In the present invention oxidizing agents are used to facilitate transdermal delivery of proteins and peptides having an Sxe2x80x94S bond across a healthy and intact skin into the blood stream to achieve a systemic effect such as, for example, in the case of insulin, the reduction of blood glucose levels. The use of transdermal delivery systems to treat systemic diseases differs form the local treatment of skin illnesses described in Rothman. The disclosure in Rothman does not relate to the delivery of proteins through skin into the blood stream. Instead, the compositions of Rothman contain proteins and oxidizers that are used to facilitate skin healing. In the present invention, however, the oxidizer prevents the reduction of disulfide bonds in the active ingredient and, thus, the structure of the active protein or peptide is retained therby enabling the systemic activity of the inventive transdermal delivery system.
Moreover, the compositions disclosed in Rothman contain a reducing agent as an essential ingredient thereof and an optional oxidizing agent. Although a therapeutic composition containing both a reducing agent and an oxidizing atent may successfully treat diseased skin according to Rothman, such a composition does not and cannot provide the desired transdermal delivery of a protein and/or peptide containing an Sxe2x80x94S bond into the blood stream, since the presence of the reducing agent required by Rothman counteracts the function of the oxidizing agent in facilitating the penetration of the active ingredient through the skin.
Thus, in preferred embodiments of the present invention there is also provided a transdermal delivery system for treating diabetes and other pathologic systemic conditions, comprising an active ingredient selected from the group consisting of peptides and proteins having an Sxe2x80x94S bond and mixtures thereof and a pharmaceutically acceptable oxidizing agent selected from the group consisting of iodine, povidine-iodine, potassium, permanganate, peroxides and silver protein and sources thereof, said system being essentially free of reducing agents and said oxidizing agent enabling and facilitating the penetration of said active ingredient through the skin layers and into the blood stream.
In initial in vivo studies, employing insulin-containing wells on rat abdominal skin, no reduction in blood glucose levels was observed. Nevertheless, insulin quantities were significantly decreased in the open wells. This discrepancy may be associated with the ability of reduced glutathione (GSH) (and of other cellular SH groups) to inactivate insulin by reducing its disulfide bond(s) [Rafter,G W (1990) Biochem. Int. 20, 817-820] followed by aggregation of several peptide molecules in the in vivo system. The opposite situation can also occur by the virtue of this invention, i.e. the effect of high levels of GSH (and other SH groups), could be overcome by topical pretreatment of the skin with oxidizing agents (such as povidone iodine, iodine alone and/or silver protein), resulting in oxidation of the cellular GSH (and other R-SH groups) to form the inactive GSSG (and/or R-SS-R). Reduced levels of the former as well as other SH containing agents prevent inactivation of insulin, enabling hormone penetration via skin into the blood stream and reduction in blood glucose levels. All tested oxidants resulted in time-dependent reduction in blood glucose levels, whereas rats without such pretreatment or treatment failed to show this phenomenon.
Povidone iodine (PI) (polyvinylpyrrolidone-iodine complex) ointment is widely used as an antiseptic agent. The successful combination of being safe and non-irritant as well as an efficient antiseptic agent was already demonstrated 40 years ago [Shelanski,H A and Shelanski,M V (1956) J. Int. Coll. Surg. 25, 727-734]. The fact that PI is commonly employed in hospitals, clinics and home use is further corroboration for its advantageous properties. Other oxidants such as silver protein (mild or strong) and permangnates, have been used for many years may also be employed in the compositions of this invention.
In the practice of this invention topical proteins such as insulin in therapeutically effective doses are incorporated into pharmaceutically acceptable carriers such as gels, ointments, solutions, paste, powder, and adhesive patch. The resulting formulations are applied to the skin of patients as many applications as needed, preferably once a day. The novel carriers contain insulin or other protein drugs that are not stable in vital skin in-vivo, thus need a protecting agent against skin biotransformation before approaching the systemic blood. The present invention comes to challenge the prior art, which considers the skin as a physical barrier to proteins such as insulin. It has been clearly proved that 5807-dalton human insulin quantitatively penetrates an excised skin in-vitro and in-vivo, but is inactivated through the in-vivo transport. This inactivation can be diminished by using oxidants such as povidone-iodine or silver protein. It has been hypothesized that these mild oxidizing agents result to a reduction in skin levels of glutathione and other reducing agents, thus preventing insulin inactivation caused by dimerization, aggregation, cleavage to two separate chains, cleavage of one Sxe2x80x94S bond, etc.
The invention is derived, in part, from the discovery that the delivery of topically-applied protein drugs having an Sxe2x80x94S bond into the blood circulation is possible only if oxidizing agents are involved before or during the application.
The present invention relates to transdermal delivery of peptide/protein drugs having an Sxe2x80x94S bond. The preferred compositions comprise a safe and effective quantities of: (a) a therapeutic protein/peptide, (b) an oxidant such as iodine or povidine-iodine, or another source of iodine or combinations of oxidants, (c) an appropriate vehicle system which may contain skin penetration enhancers selected from those known in the art (see, e.g., Ghosh, T. K., Pfister, W. R., and Yum, S. I., Transdermal and Topical Drug Delivery Systems,1997, Interpharm Press, Inc., pp.357-509).
Effective levels of protein drugs are delivered by the novel compositions. An xe2x80x9ceffectivexe2x80x9d level is meant that a concentration of a drug is high enough to be effective in treating the condition in which the drug has been designed to treat. Examples of protein drugs are: insulin, alpha-, beta-, and gamma-interferon, human growth hormone, alpha- and beta-1-transforming growth factor, granulocyte colony stimulating factor (G-CSF), granulocyte macrophage colony stimulating factor (G-MCSF), parathyroid hormone (PTH), human or salmon calcitonin, glucagon, somatostatin, vasoactive intestinal peptide (VIP), and LHRH analogs.
As mentioned above, povidone-iodine, iodine and sources thereof as well as silver protein are preferably selected as safe and effective xe2x80x9cskin-protein-stabilizerxe2x80x9d in a pretreatment procedure or in treatment composition at concentrations of 0.01% to 80%.
The therapeutic proteins and its protectors/stabilizers in-vivo, can be applied as a topical formulation such as cream (o/w or w/o), ointment, film-forming liquid spray, or gel using occlusive or non-occlusive dressings. A composition in an appropriate polymeric patch is preferable as transdermal protein delivery, and it consists of compatible adhesive polymers known in the art. The polymer can be selected from the group of acrylic polymers, cellulosic polymers, polyurethanes, polylactic/polyglycolic acids, polyamino acids, polysaccharides, polyurea, polyvinyl alcohol, polyvinylpyrrolidone (povidone), and natural proteins.
A transdermal patch can consist several layers: in the inner side a peelable plastic cover will protect the drug layer containing the adhesive polymer, plasticizer, the oxidizing agents, penetration enhancers and other excipients. The outer layers (i.e., the external layers) are designated to protect the drug from diffusion outward and to stick the patch by its margins to the skin, so the drug layer is occluded from all sides except the skin side where it is in close contact (see, e.g., Chien, Y. W., Transdermal Controlled Systemic Medications, 1987, Marcel and Decker, pp. 93-120, 365-378).
While the invention will now be described in connection with certain preferred embodiments in the following examples and with reference to the accompanying figures so that aspects thereof may be more fully understood and appreciated, it is not intended to limit the invention to these particular embodiments. On the contrary, it is intended to cover all alternatives, modifications and equivalents as may be included within the scope of the invention as defined by the appended claims. Thus, the following examples, which include preferred embodiments will serve to illustrate the practice of this invention, it being understood that the particulars shown are by way of example and for purposes of illustrative discussion of preferred embodiments of the present invention only and are presented in the cause of providing what is believed to be the most useful and readily understood description of the formulations procedures as well as of the principles and conceptual aspects of the invention.